Squamous cell carcinoma of the head and neck (SCCHN) is a significant public health concern affecting over 50,000 patients annually in the US. Recurrence rates in HPV-negative SCCHN are approximately 50%, while the median overall survival for patients with recurrent/metastatic disease is approximately 1 year, thus novel therapies are urgently needed. Protein lysine methyltransferases (PKMTs) are a class of histone modifiers that are mostly known to regulate the histone epigenome through methylation of specific histone lysine residues. With the advent of the Cancer Genome Atlas (TCGA), PMTs were found to be mutated, amplified or overexpressed in SCCHN. The role of NSD3 in SCCHN: The PKMT NSD3 was found to be among the top 10 most frequently amplified genes in SCCHN with a significantly recurrent amplification at the 8p11.23 region, where the gene is located, in approximately 10% of SCCHN tumors, implying its function as an oncogene. We have recently shown that NSD3 promotes oncogenesis through induction of H3K36 di-methylation and transcriptional upregulation of the critical cell cycle regulators CDC6 and CDK2. Our findings also revealed that NSD3 directly binds and mono-methylates EGFR, enhancing its interaction with proliferating cell nuclear antigen (PCNA) and promoting cell cycle progression in SCCHN cells. Despite the importance of NSD3 in SCCHN oncogenesis, the genome-wide distribution of NSD3 in head and neck cancer cells, and the mechanisms that direct it to specific genomic loci are unknown. This project will elucidate the genome-wide distribution of the short and the long isoforms of NSD3 in head and neck cancer cell lines, it will assess proteins interacting with each of the isoforms and evaluate post-translational modifications and their functional impact, including the phosphorylation status (probably mediated by EGFR) of each of these isoforms. Major Activities: To answer the questions above, our experimental approach will include chromatin immunoprecipitation followed by DNA sequencing, RNA sequencing in stably transfected SCCHN cell lines expressing the short or the long isoform of NSD3. The role of SMYD3 in SCCHN: In this project, we will assess the immunomodulatory and oncogenic functions of SMYD3 in SCCHN. To this purpose, we will be conducting CHIP-seq, ATAC-seq and RNA-seq to identify direct downstream targets of SMYD3, assess if it affects chromatin condensation and the enhancer activation status in SCCHN cells, as well as mouse experiments to evaluate the growth and tumor immunomodulatory effects of SMYD3.